There are mainly two methods for generating RNA molecules. They are 1) solution-phase based in vitro transcription using T7 phage RNA polymerase (hereafter referred as solution-based T7 transcription), and 2) solid-phase based chemical synthesis from phosphoramidites using a synthesizer. Solution-based T7 transcription has widely been used to generate transcript RNAs as large as several kilobases in large quantity, but is not applicable for generating specifically labeled RNAs. Chemical synthesis is used to obtain small RNAs (up to 40-60 bases long) in minute quantity, limited by an intrinsically low coupling rate compared to the DNA counterpart. It is possible to use the chemical synthesis method to generate selective labeled small RNAs, provided that all labeling reagents are commercially available. But specifically labeling sizable RNAs using the chemical synthesis is not practical, due an extremely low efficiency and prohibitively high cost.
Selective labeling of specific residue(s) and/or specific region(s) of RNAs (SLOR) is now possible due to the development of the SLOR technology. The same type of labeling has not yet become practical even for DNA and proteins. Provided herein is a hybrid solid-liquid phase enzymatic method that allows specific labeling at designated residue(s) and/or segment(s) of an RNA. Selected residues can be specifically labeled with stable isotopes such as 13C/15N, or with fluorophores such as Cy3 and Cy5 rCTP, rUTP, or rNTP derivatives. The efficiency of the method is similar to that using the solution-phase T7 transcription.